title: Foxc2 regulates osteogenesis and angiogenesis of bone marrow mesenchymal stem cells.
abstract: BACKGROUND
  The Forkhead/Fox transcription factor Foxc2 is a critical regulator of osteogenesis and angiogenesis of cells. Bone marrow mesenchymal stem cells (BMSCs) have the capacity to differentiate into osteoblasts, chondrocytes, adipocytes, myocytes and fibroblasts. The present study investigates the role of Foxc2 overexpression in osteogenesis and angiogenesis of BMSCs in vitro.
  
  
  METHODS
  BMSCs were isolated from SD rat femurs and tibias, and characterized by cell surface antigen identification and osteoblasts and adipocytes differentiation. The cells were transfected with lentiviral Foxc2 (Lv-Foxc2) or green fluorescent protein (Lv-GFP). Seventy hours later, Foxc2 expression was observed using real time-PCR and Western blot. The transfected cells were stained with Alizarin red S or alkaline phosphatase (ALP) after osteogenic induction. Meanwhile, the expression levels of osteocalcin (OCN), Runt-related transcription factor 2 (Runx2), vascular endothelial growth factor (VEGF) and platelet-derived growth factor-β (PDGF-β) were measured by real time-PCR, Western blot and immunostaining.
  
  
  RESULTS
  Results of cell characterization showed that the cells were positive to CD44 (99.56%) and negative to CD34 (0.44%), and could differentiate into osteoblasts and adipocytes. Foxc2 overexpression not only increased the numbers of mineralized nodes and ALP activity, but also enhanced the expressions of Runx2, OCN, VEGF and PDGF-β in transfected BMSCs after osteogenic induction. The effects of Foxc2 on osteogenesis and angiogenesis were significantly different between Lv-Foxc2 transfected BMSCs and Lv-GFP transfected BMSCs (P<0.05). In addition, the MAPK-specific inhibitors, PD98059 and LY294002, blocked the Foxc2-induced regulation of BMSC differentiation.
  
  
  CONCLUSIONS
  Foxc2 gene is successfully transfected into BMSCs with stable and high expression. The overexpression of Foxc2 acts on BMSCs to stimulate osteogenesis and angiogenesis. The effect of Foxc2 on angiogenesis of the cells is mediated via activating PI3K and ERK.
authors.affiliation: Department of Orthopedics, The Second Affilliated Hospital of Xi'an Jiaotong University, Xiwu Road, Xi'an, Shaanxi Province 710004, China.
authors.forename: Wulin
authors.initials: W
authors.lastname: You
authors.forename: Hongwei
authors.initials: H
authors.lastname: Gao
authors.forename: Lihong
authors.initials: L
authors.lastname: Fan
authors.forename: Dapeng
authors.initials: D
authors.lastname: Duan
authors.forename: Chunsheng
authors.initials: C
authors.lastname: Wang
authors.forename: Kunzheng
authors.initials: K
authors.lastname: Wang
mesh_terms: D017667:Adipocytes / Q000166:cytology / Q000378:metabolism; D043925:Angiogenesis Inducing Agents; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D048049:Extracellular Signal-Regulated MAP Kinases / Q000096:biosynthesis; D051858:Forkhead Transcription Factors / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D018919:Neovascularization, Physiologic / Q000235:genetics*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics*; D019869:Phosphatidylinositol 3-Kinases / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection
chemical_list: D043925:Angiogenesis Inducing Agents; D015415:Biomarkers; D051858:Forkhead Transcription Factors; C082078:mesenchyme fork head 1 protein; D019869:Phosphatidylinositol 3-Kinases; D048049:Extracellular Signal-Regulated MAP Kinases